The essential anticoagulant protein antithrombin III (ATIII) uses a serpin suicide substrate mechanism to form inhibitory complexes with clotting enzymes. However, native circulating ATIII is an inefficient proteinase inhibitor due to partial insertion of its reactive loop in its central A beta-sheet. For full activation, cofactor heparin must bind ATIII and induce a protein conformational change that leads to reactive loop expulsion and about 300x increase in the fXa inhibition rate. The mechanism of ATIII activation by heparin is reasonably well understood at the first stage of cofactor binding to the inhibitor and the final stage of reactive loop expulsion. In contrast, much less is known about intermediate steps responsible for propagation of the activating conformational change from the pentasaccharide-binding site to the reactive loop. The goals of this work are to identify ATIII structural elements and interactions that mediate conformational change transmission, and to develop a better understanding of this pharmaceutically important allosteric activation mechanism. Our working model proposes that 3 prongs of conformational change radiate from the pentasaccharide-ATIII interface. Prong-1 leads to helix P (hP) formation and then converges with prong-2 (which is transmitted through the N-terminal polypeptide) to restructure the hE arm and rotate Y166. Y166 rotation promotes sheet-A closure at the equator of the inhibitor. Sheet A closure at the reactive loop pole is similarly promoted by prong-3-mediated Y131 rotation. (Prong 3 is relayed through hA, sB and hD following ATIII binding to the pentasaccharide non-reducing end.) As strands 2 and 3 of sheet-A move away from hD and hE, they pass under the hF arm which with a network of A-sheet residues directly stabilizes the inserted reactive loop and partially opened A-sheet of the native conformation. Tyrosine rotation-driven movement of s2A/s3A under the hF arm disrupts this native network and frees its components to engage in different linkages that stabilize the P14-expelled closed A-sheet activated molecule. The model for heparin dependent conformational change propagation in ATIII will be evaluated and refined by disrupting hypothesized, critical structural interactions and determining how such changes affect heparin affinity and binding kinetics, allosteric activation of fXa inhibition and the molecular structures of the mutants.